The formation of neural circuits relies on activity-dependent and independent mechanisms during development. Before sensory systems provide input to the nervous system, networks such as the spinal cord, retina and hippocampus display spontaneous, rhythmic bursts of action potentials. The spatiotemporal patterns of this spontaneous activity have been shown to play a significant role in the development of the visual system. Little is known, however, about how patterns of neural activity in the spinal cord contribute to the maturation of motor circuitry. Uncovering the influence of spontaneous activity on the maturation of the central pattern generator (CPG) would provide elementary insights into developmental mechanisms supporting basic behaviors like walking and swimming. This knowledge would inform regenerative technologies that seek to stimulate the growth and integration of new neural networks into the spinal cord by identifying the activity requirements for analogous developmental processes. Global patterns of spontaneous activity in the spinal cord have been shown to be locomotor-like in vertebrate models such as the rat and chick, with synchronization in ipsilateral regions of the spinal cord and alternation left and right. The goal of our research plan is to establish how these coordinated patterns of spontaneous activity are acquired and what role they play in the formation of a functional motor circuit. We have chosen the zebrafish as a model system. To accomplish our goals, we will: 1) characterize the spatiotemporal patterns of spontaneous activity in the zebrafish spinal cord, 2) identify the cell types that mediate the coordination of this activity, and 3) determine the role that these patterns of activity play in the formation of the CPG and the generation of basic behaviors. We will take full advantage of the transparency and genetic accessibility of the zebrafish and apply genetically-encoded optical tools for our study. We will monitor the spatiotemporal patterns of spontaneous activity in defined cell populations with the genetically-encoded calcium indicator GCaMP. To identify cell types mediating coordinated activity, we will use the light-driven chloride pump Halorhodopsin and the photosensitizer KillerRed to perform acute and chronic optical lesions of defined cell types. Finally, we will alter the patterns of spontaneous activity with Halorhodopsin and the light-gated cation channel Channelrhodopsin and observe consequences on the development of the CPG by assaying simple behaviors.